Project summary/Abstract The long-term goal of this project is to improve therapeutic options for patients with cholangiocarcinoma by better understanding the pathways that drive disease and increase resistance to therapy. Worldwide, liver cancer is now second only to lung cancer for cancer- related deaths. Cholangiocarcinoma is a primary liver cancer with rising incidence and few effective treatment options. The current project will investigate canonical and non-canonical fibroblast growth factor receptor-4 (FGFR4) signaling in cholangiocarcinoma initiation and progression. Preliminary data demonstrated that cholangiocarcinoma cells exhibited autocrine FGFR4 signaling through expression and secretion of the ligand FGF19. Inhibition of this pathway increased apoptosis and decreased cell growth with effects at the G1/S checkpoint, S phase duration, and mitosis. FGFR4 is strongly expressed in tumor tissue compared to normal bile ducts. FGFR inhibition markedly reduced tumor size in rats and knockdown of FGFR4 prevented subcutaneous tumor formation. We have identified a novel proteolytic cleavage product of the full-length receptor that is composed of the intracellular domain (R4- ICD). R4-ICD contains the C-terminal tyrosine kinase domain, activated AKT, and prevented apoptosis. The central hypothesis of this proposal is that signaling through FGFR4 and R4-ICD drives cholangiocarcinoma initiation, progression, and chemoresistance. Experiments in aim 1 will determine how malignant features are promoted by R4-ICD and FGFR4. Aim 2 will identify tumor-initiating mechanisms of FGFR4 and R4-ICD. The third aim will demonstrate the role FGFR4 and R4-ICD in chemotherapy resistance. Successful completion of this project will increase our understanding of how FGFR4 promotes cholangiocarcinoma progression and resistance to therapy, define new biology of R4-ICD, and test treatment strategies that inhibit FGFR4 processing or kinase activity.